The process of aging affects the function of T-lymphocytes as exemplified by decreased cellular proliferation in response to TCR and CD28 co-ligation. This decline is in part due to decreased IL-2 production, which may be causally related to decreased activation of NFAT and AP-1 response elements in the IL-2 promoter. To this end, TCR alone or TCR plus CD28 co-ligation induces different MAP kinase cascades which regulate NFAT/AP-1 response elements in the IL-2 promoter. In addition to proliferation, senescent T-cells also exhibit defects in apoptosis which may be causally related to decreased Fas (CD95) expression and function. Insofar as Fas induces the Jun Kinase cascade, which has a pro-apoptotic effect, it is possible that altered regulation of this MAPK family may be involved in apoptosis dysregulation in aged T-cells. As a unifying mechanism, we propose an integrating role for the ERK and JNK cascades which determines cellular AP-1 composition. Moreover, we propose that the dynamic equilibrium which exists between these cascades governs whether a T-cell will proliferate or undergo apoptosis. Our long-term goal is to understand the molecular mechanisms which lead to decreased T- cells function in aged individuals. In aim 1 we will dissect the roles of the MAPK pathways in the activation of AP-1 response elements in the IL-2 promoter as it varies with age. In T-cell lines, we will determine what effect stable, tetracycline-regulated expression of various combinations of dominant active or dominant negative MAPK regulators has on NFAT/AP-1 response elements in the IL-2 promoter. In splenic T-cells from young and aged mice, we will determine whether altered regulation of ERK or JNK pathways in response to TCR and CD28 co-engagement impact on the expression of AP-1 proteins and binding to individual NFAT/AP-1 response elements in the murine IL-2 promoter. In aim 2 we will dissect the role of the MAPK pathways in the control of apoptosis in human T-cell lines as well splenic T-lymphocytes from young and aged mice. In T-cell lines, stably transfected with various combinations of inducible dominant negative or active regulators, of the ERK and JNK cascades, we will study the molecular mechanisms by which these pathways regulate induction of apoptosis, including possible effects on the CD95 promoter. We will compare various pro-apoptotic stimuli on MAPK activation in young and aged murine T-lymphocytes. We will use T-cell lines to determine whether these MAPK cascades regulate Bcl-1, Bcl-xl, Bax expression. We will determine whether there is altered expression of these proteins in T-cells from aged mice.